Arginase is an enzyme which is associated with liver cells and its presence in serum is generally regarded as an indication of liver disease.
Accordingly, various means have been attempted for measuring arginase levels in animals. However, attempts to assay arginase per se have proved unreliable because it is an enzyme which can be inactivated or lost during isolation; therefore, any procedure which relies upon a direct measurement cannot assure that the results accurately reflect the concentration of arginase in the original sample.
Consequently, the search for a reliable test has taken other approaches. One of these consists of measuring the products which arginase is known to produce. It is known, for example, that arginase hydrolyzes serum arginine to urea and ornithine and, as a result, considerable effort has been expended in developing an assay which can reliably measure the ornithine and urea produced from a given amount of L-arginine: ##STR1## However, efforts at measuring the urea product of this reaction are impractical because of the residual urea normally present in serum.
Therefore, various methods have been attempted at measuring the ornithine product of that reaction. Unfortunately, however, direct spectrophotometric measurements have proved unsuccessful because the spectral curve of ornithine is so similar to that of arginine that it cannot serve as a reliable basis for routine testing.
One method which has proved moderately successful is the procedure of W. Roman and J. Ruys reported in Clinical Enzymology, Vol. 2: pages 121-128 (1970). This procedure consists of treating an arginase-containing serum sample with L-arginine to produce urea and ornithine. The resulting sample is subsequently treated with trichloroacetic acid and a ninhydrin indicator in acidic medium to produce a colored ornithine-ninhydrin complex which can be spectrophotometrically measured.
By measuring the optical density of both the test sample and a blank in which no enzymatic reaction has taken place, the concentration of ornithine in the test material can be determined. From this measurement an assay for arginase can be computed.
Although this method is reliable, it is cumbersome and cannot be used with the efficiency which is generally required in large scale testing and routine clinical studies.
One difficulty lies in the fact that a precipitation and centrifugation step must be conducted in order to obtain a clear supernatant liquid suitable for spectrophotometric comparison.
One other drawback is the need to utilize trichloroacetic acid as one of the reaction ingredients. In addition to its very corrosive and caustic properties, trichloroacetic acid, even of analytical grade, generally contains side products which interfere with the ornithine assay.